The invention generally relates to platinum compounds, which may be used for detectably labeling biological target molecules. More particularly, the invention relates to platinum-based linker compounds, labeling compounds, and detectably labeled molecular probes and methods of using such compounds for the detection and localization of biological substances of interest.
Platinum (coordination) compounds have been considered interesting molecules for a very long time. For a review of some of these compounds and their uses we refer to Reedijk et al., J. Struct. Bonding, 67: 53-72 (1987). This article describes the anti-tumor compound cis-Pt (NH.sub.3).sub.2 Cl.sub.2, which compound has a high affinity for certain biological or bio-organic molecules, including amongst others, proteins and DNA molecules. In particular, it appears that such compounds have a marked affinity for the N.sup.7 -nitrogen atom in the purine bases guanine and adenine, as well as for sulfur groups in macromolecules.
By dissociation of the two chlorine ligands in these compounds, two reactive sites arise, with which such platinum compounds can cross-link within or between molecules. For example, such compounds are known to cross-link between two neighboring guanine and/or adenine bases in the same or opposite DNA strands, thereby inhibiting the replication of the DNA molecules. The application of cis-platinum (cisplatin) as an anti-tumor drug (cytostaticum) is based on this mechanism.
Other applications of the cross-linking capacity of such compounds have been explored. For example, U.S. Pat. No. 4,490,543 discloses platinum-based radiation sensitizers which are said to be derivatives of cisplatin. These sensitizers contain two reactive groups and, accordingly, cross-link biological molecules such as DNA.
In contrast, monochlorinated platinum compounds like Pt(dien)Cl appear to keep their DNA affinity, but they do not form cross-links and interfere only slightly with the base pairing of complementary DNA strands. As such, these compounds have no anti-tumor activity.
According to U.S. Pat. No. 4,711,955, non-radioactive nucleic acid labeling techniques are preferred in present medical-biological practice, especially diagnostic practice. The presently applied known non-radioactive labeling techniques for DNA and RNA are globally to be divided in two categories:
1. Labeling which proceeds via enzymatic or organic synthetic routes; for instance biotin, bromodeoxyuridine (BrdU), digoxigenin, fluorescein and peroxidase; PA0 2. Labeling by direct chemical coupling, like photobiotin, AAF, mercury, and sulfone groups. The '955 document discloses one such labeling method, but does not provide any information concerning use of platinum-based compounds for this purpose. PA0 1. Direct and almost instantaneous labeling of macromolecules without necessity of enzymatic or organo-synthetic procedures; PA0 2. one-step purification of labeled molecules by means of a simple routine technique; PA0 3. Direct and/or indirect detection of labeled molecules by way of almost all known (microscopic) techniques.
Application of detectable labels to biomolecules has been typically accompanied by a number of problems. Such problems are a result of several factors, including the complexity of previous labeling procedures, limitations on the length of the synthetic oligonucleotides which have been desired to be labeled, the necessity of using health-injuring compounds, and a lack of stability of the label when it has been bound to the nucleic acid.
Given these limitations in the conventional methods of labeling bio-organic target molecules, it is desirable to provide labeling method which overcomes these problems. Probe compounds are desired which can accommodate ready attachment of a large variety of detectable markers. It is also desirable to provide detectably labeled probe compounds which can quickly and easily identify specific bio-organic target molecules such as nucleic acids and proteins. In addition, it would be desirable to provide detectably-labeled probe compounds which would specifically interact with particular bio-organic target molecules, but which do not substantially interfere with their function.
The present invention now provides platinum-containing compounds, in the application of which the above-mentioned disadvantages are effectively removed.